Saccharides enhance iron bioavailability to Southern Ocean phytoplankton
Edited by David M. Karl, University of Hawaii, Honolulu, HI, and approved November 19, 2010 (received for review July 26, 2010)
Commentary
January 5, 2011
Abstract
Iron limits primary productivity in vast regions of the ocean. Given that marine phytoplankton contribute up to 40% of global biological carbon fixation, it is important to understand what parameters control the availability of iron (iron bioavailability) to these organisms. Most studies on iron bioavailability have focused on the role of siderophores; however, eukaryotic phytoplankton do not produce or release siderophores. Here, we report on the pivotal role of saccharides—which may act like an organic ligand—in enhancing iron bioavailability to a Southern Ocean cultured diatom, a prymnesiophyte, as well as to natural populations of eukaryotic phytoplankton. Addition of a monosaccharide (>2 nM of glucuronic acid, GLU) to natural planktonic assemblages from both the polar front and subantarctic zones resulted in an increase in iron bioavailability for eukaryotic phytoplankton, relative to bacterioplankton. The enhanced iron bioavailability observed for several groups of eukaryotic phytoplankton (i.e., cultured and natural populations) using three saccharides, suggests it is a common phenomenon. Increased iron bioavailability resulted from the combination of saccharides forming highly bioavailable organic associations with iron and increasing iron solubility, mainly as colloidal iron. As saccharides are ubiquitous, present at nanomolar to micromolar concentrations, and produced by biota in surface waters, they also satisfy the prerequisites to be important constituents of the poorly defined “ligand soup,” known to weakly bind iron. Our findings point to an additional type of organic ligand, controlling iron bioavailability to eukaryotic phytoplankton—a key unknown in iron biogeochemistry.
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Dissolved iron supply sets the rate of primary production and nutrient utilization in up to 40% of open-ocean waters, including the Southern Ocean (1, 2). Because of the complex and dynamic interplay between iron chemistry, photochemistry, and biological processes (Fig. S1), the mechanism(s) controlling iron bioavailability to oceanic biota remain poorly understood. Iron bioavailability is influenced by photochemistry (3), chemical speciation (4, 5), biological cycling (6), and uptake strategies (7–9). As a result, different microorganisms, each with its own specific iron requirement, iron uptake system(s), and biological adaptability (7, 10, 11), will have access to different pools of bioavailable iron under conditions of identical iron chemistry.
In marine systems, virtually all of the iron is bound to organic ligands (12, 13); hence, organic complexation is the main regulator of both iron reactivity and bioavailability. Both experimental (4, 14) and modeling (15) observations agree that organic complexation is necessary to maintain iron solubility and bioavailability in the ocean. In oceanic waters, two classes of organic ligands (L1 and L2) have been identified on the basis of their binding affinities for iron (12). It is widely accepted, from comparable conditional stability constants, that siderophores produced by bacterioplankton (i.e., heterotrophic and autotrophic bacteria) belong to the strongest class (L1) of organic ligands. However, a photoactive siderophore that belongs to the L2 class has now been reported (3). Moreover, uncertainties remain about the nature of organic ligands that bind iron (12, 16). Ligands too weak to fall within the analytical window of the competitive ligand exchange technique are also not detected (17); thus, their importance for iron biogeochemistry is likely to be underestimated. There is no clear evidence to date that eukaryotic phytoplankton produce or use siderophores. Nevertheless, whatever their source, the presence of photolabile siderophores could improve iron biavailability through the indirect photoreduction of iron(III) to iron(II). The single instance reported so far has been the iron-binding siderophore, vibrioferrin, increasing iron bioavailability to a cultured dinoflagellate (3). Therefore other types of ligands, rather than siderophores, might be controlling iron bioavailability to eukaryotic phytoplankton.
Saccharides are poorly defined, polydisperse, and polyfunctional compounds having “weak” affinities for cationic metals (17–19). Gluconic and domoic acids both form weak iron complexes that fall within the L2 class (log KFe′L 8.7 and 8.8 M−1, respectively) (17, 20). Unlike siderophores, reported at picomolar levels (21), saccharides occur in relatively high concentrations in surface waters (nanomolar to micromolar) (22), giving them the potential to outcompete the L1 class for iron binding—a critical step toward defining iron bioavailability (20). Saccharides represent 3–50% of dissolved and colloidal organic matter (18, 23, 24) and are typically present at 0.2–500 nM in the Pacific Ocean (22), representing up to 20–30 μM of carbon (23). Polysaccharides are more abundant than monosaccharides in surface water, comprising up to 70% of total saccharides (23). The concentration of polysaccharides decreases with depth, suggesting that polysaccharides are reactive components in surface water (23). Saccharides are ubiquitous and important constituents of marine gels, transparent exopolymeric particles, and dissolved organic carbon (22, 24).
Most marine microorganisms, including eukaryotic phytoplankton and heterotrophic and autotrophic bacteria, synthesize saccharides that are either internal energy stores or secreted as exopolysaccharides (EPS) (24–29). Storage saccharides are metabolism dependent, responsive to iron supply (30), and released by both cell lysis and grazing—two iron-recycling pathways (6). Indeed, saccharides can influence trace metal chemistry and cycling (31), two parameters that define bioavailability (Fig. S1). Evidence exists that saccharides might be important for iron chemistry and bioavailability; however, no prior studies have attempted to develop a mechanistic understanding of the effect of saccharides on iron bioavailability. The excretion of domoic acid by Pseudonitzschia was increased under conditions of iron limitation (20, 32). Saccharides can also increase the growth of Southern Ocean eukaryotic phytoplankton (33) and favor iron reduction, which releases more soluble iron(II) (34).
Saccharides vary in their potential to bind iron. Uronic acids, for example, are likely to be significant for iron biogeochemistry; they are a major constituent of the natural saccharides found in surface waters, as well as in the EPS excreted by eukaryotic phytoplankton and heterotrophic and autotrophic bacteria (24, 26–29, 35, 36). The carboxylic groups of uronic acids are known to bind transition metals, including iron(III) (19, 35, 37). To understand the underlying mechanisms by which saccharides (including uronic acids) alter iron bioavailability to eukaryotic phytoplankton, we measured the intracellular iron uptake, chemical complexation, and solubility with and without the addition of organic ligands (two saccharides, one bacterial EPS, and one siderophore). Here, we present compelling evidence that saccharides can significantly increase the iron uptake rate of eukaryotic phytoplankton for both natural assemblages and isolates from the Southern Ocean. Hence, we propose that saccharides are important organic compounds controlling iron bioavailability to eukaryotic phytoplankton in iron-deficient waters, and in particular for high-nutrient, low-chlorophyll (HNLC) oceanic regions.
Results
Iron Bioavailability to Natural Planktonic Assemblages.
Under iron deficiency, the bioavailable iron pool is defined here as any iron form that can accumulate within microorganisms to support metabolic reactions and their growth (Fig. S1). Bioavailability, therefore, is related to the intracellular iron uptake rate (38). In our study, we selected the uronic acid, glucuronic acid (GLU), as it is a major constituent in the EPS of heterotrophic and autotrophic bacteria, diatoms, Emiliania huxleyi, and Phaeocystis (26–29, 35, 36). The siderophore desferrioxamine B (DFB, a natural siderophore from a soil bacterium) was selected as a contrasting ligand, because it forms strong organic complexes with iron (13, 17) and generally decreases iron bioavailability to eukaryotic phytoplankton (38).
DFB decreased iron bioavailability to both resident bacterioplankton and eukaryotic plankton at the polar front (PF) (Fig. 1A and Table S1) and in the subantarctic zone (SAZ) (Fig. 1C). The effect of DFB on decreasing the iron uptake rate was threefold greater for eukaryotic plankton than for bacterioplankton. In the PF, the addition of GLU, a monosaccharide, decreased the iron uptake rate of bacterioplankton to a degree comparable to that observed for DFB. However, GLU increased the iron uptake rate to eukaryotic phytoplankton by 2.3-fold (Fig. 1B). In the SAZ, the iron uptake rate of both bacterioplankton and eukaryotic plankton was increased in the presence of ≤2 nM of GLU (Fig. 1D). This site-specific effect of GLU observed for bacterioplankton may be due to the presence of autotrophic bacteria representing 10% of total bacterioplankton found at the SAZ site compared with 0.01% at the PF site (Table S1). Despite different dominating eukaryotic plankton (diatoms at PF; dinoflagellates at SAZ; Table S1), similar trends were evident for both SAZ and PF sites. For GLU ≥2 nM, the iron uptake rate was enhanced threefold more for eukaryotic plankton than for bacterioplankton (Fig. 1D). Although there were comparable concentrations of dissolved iron and chlorophyll a (Chl a) in the SAZ and in the PF (Table S1), the iron uptake by eukaryotic plankton in the SAZ was 5.6-fold greater than in the PF. Furthermore, with a 3.7-fold lower bacterioplankton biomass at the PF compared with the SAZ (Table S1), the iron uptake by bacterioplankton at the PF was 0.7-fold less compared with the SAZ (Fig. 1, no organic ligand added). This suggested a difference in the degree of iron bioavailability between the planktonic communities at the PF and SAZ sites.
Fig. 1.
Iron Bioavailability to Phytoplankton Isolates.
Results obtained in the field were confirmed by laboratory experiments using axenic eukaryotic phytoplankton isolates from the Southern Ocean. In Antarctic filtered seawater (Fig. 2A), for both Chaetoceros sp. and Phaeocystis sp., iron uptake rates were significantly increased in the presence of GLU and dextran (DEX, a polysaccharide) compared with iron uptake rates in the presence of inorganic iron (P = 0.005–0.040). In artificial seawater (Fig. 2B), iron uptake rates were significantly increased in the presence of GLU for Chaetoceros sp. (P = 0.026) and in presence of DEX for both isolates (P = 0.001–0.003). The addition of DFB significantly decreased the iron uptake rates for both isolates (P = 0.000–0.003; Fig. 2 A and B). Iron bioavailability was increased in the presence of GLU for both natural populations of eukaryotic phytoplankton and laboratory axenic isolates. This observation demonstrated that the enhancement was independent of bacterial activity, such as iron remineralization and GLU as a potential carbon source for heterotrophic bacteria. In addition, comparable trends for both synthetic inorganic and natural ocean waters (Fig. 2 A and B) suggested that the enhanced iron bioavailability was mainly due to saccharides rather than to a combined effect of saccharides and other natural ligands. Because DEX is too large to be directly taken up by eukaryotic phytoplankton, the enhanced iron uptake rate is likely to be related to an iron uptake pathway rather than to a saccharide uptake pathway.
Fig. 2.
The intracellular uptake rate constant (kupt, Table 1) is a measure of the bioavailability of iron associated with saccharides, which relates directly to the biological transport affinity of the chemical forms of iron present (39). According to kupt in the absence of organic ligands, Phaeocystis sp. takes up iron threefold faster than Chaetoceros sp. (Table 1). The comparison of kupt in the presence of inorganic iron and in the presence of saccharides provided a measure of the bioavailability of iron associated with saccharides. For Chaetoceros sp., no difference in kupt was observed in the presence of 5 nM of GLU and DEX, indicating no statistical difference in iron bioavailability (Table 1). In the presence of 1 nM of EPS, kupt was significantly lower, indicating a decreased iron bioavailability (Table 1). Considering the 90–98% decrease in the iron uptake rates in the presence of DFB (Fig. 2), iron associated with the EPS was still significantly bioavailable to Chaetoceros sp. and Phaeocystis sp.
Table 1.
| kupt (h−1) (×10−5) | Bioavailability, % | |||
|---|---|---|---|---|
| Chaetoceros sp. | Phaeocystis sp. | Chaetoceros sp. | Phaeocystis sp. | |
| No organic ligand | 18.3 ± 0.81 | 55.7 ± 0.35 | 100 ± 7 | 100 ± 1 |
| EPS, 1 nM | 5.00 ± 0.25 (0.004) | 19.6 ± 0.27 (0.0001) | 28 ± 2 | 35 ± 1 |
| GLU, 5 nM | 20.6 ± 1.28 (0.266) | ND | 113 ± 10 | ND |
| DEX, 5 nM | 15.3 ± 1.17 (0.171) | ND | 84 ± 9 | ND |
kupt, intracellular uptake rate constant for Antarctic eukaryotic phytoplankton incubated in artificial seawater. Average kupt (n = 2) ± interval/2 and average relative bioavailability (n = 4) ± SD are presented. ND, not determined. The kupt is determined by the linear regression of iron uptake rates at increasing iron concentrations (ligand concentration is constant). A lower value indicates a decrease in iron bioavailability measured at the level of the biological iron transporters. The numbers in parentheses refer to the P value from the paired Student's t test, comparing the kupt in the absence and in the presence of organic ligands. The average relative bioavailability for each ligand is calculated by comparing the kupt in the absence and in the presence of the ligand.
Iron Chemical Speciation.
Iron bioavailability is related to iron chemistry (refs. 4, 9; Fig. S1) and chemically labile iron complexes are thought to be more bioavailable (39). The competitive ligand exchange–adsorptive cathodic stripping voltammetry technique (CLE–AdCSV) in synthetic seawater was used to demonstrate the association of iron with organic ligands (17). The presence of DFB resulted in strong complexes reducing iron lability (Fig. 3A), whereas GLU did not decrease iron lability. This observation suggested the formation of a weak Fe-GLU association, outside the analytical detection window (Fig. 3A). The iron content of the EPS was 2.15 ± 0.28 mole total iron per mole of EPS. Of this total iron, only ∼38% was labile (0.82 ± 0.04 mol per mole of EPS, Fig. 3B). The inability to saturate the EPS with iron (see also results on iron solubility, Table S2), prevented the determination of the conditional stability constant of iron associated with EPS (13, 17).
Fig. 3.
An electrochemical method, recently developed to measure Suwannee River fulvic acid (SRFA)-like compounds (40) was used to measure organic compounds likely to influence iron chemical speciation at both the PF and in the SAZ. Here, 7.4 ppb (PF) and 33.6 ppb (SAZ) SRFA equivalents were measured at the depth of Chl a maximum (Table S1). However, this analytical approach is not specific to humic substances; other types of compounds differing from humics, such as glutathione, can be detected (40). EPS was also detected by this technique with 22 nM of EPS (37.4 ppm of EPS) corresponding to 37.9 ppb of SRFA equivalent.
Iron Solubility.
Saccharides increase iron solubility due to their associations with iron. For example, in the presence of 1 nM of EPS and 23.8 nM of total iron in synthetic seawater, 9.8 nM of iron was within the dissolved phase (with two-thirds of this being in the colloidal phase, Table S2). Here, EPS stabilized iron in the colloidal phase, improving its retention within the dissolved phase. Stabilization of iron in the soluble and colloidal phases was also seen for GLU and DEX in both synthetic and filtered Antarctic seawater (Table S2).
Discussion
The intracellular iron uptake rates in the presence of saccharides (GLU and DEX) were higher than for inorganic iron at identical total iron concentrations. This trend was evident for eukaryotic phytoplankton, both resident populations from two Southern Ocean locations, and two axenic isolates. Therefore, this effect is unlikely to be related to site-specific biological iron requirements or iron uptake strategies. Hence, it points to a generic (i.e., taxa independent) role for saccharides in enhancing iron bioavailability to eukaryotic phytoplankton. Saccharides with different functional groups and sizes (GLU, DEX, galacturonic acid, and alginic acid; this study, ref. 38) all enhanced the bioavailability of iron to a range of Southern Ocean eukaryotic phytoplankton. This observation may thus be applicable to other saccharides and phytoplankton groups.
In the Southern Ocean, the growth of eukaryotic phytoplankton is largely limited by iron (1) and both eukaryotic phytoplankton and bacterioplankton are competing for this micronutrient. The contrasting results obtained for natural eukaryotic phytoplankton and bacterioplankton communities in the presence of GLU suggest that saccharide associations with iron might represent an advantage for iron uptake and growth of eukaryotic phytoplankton in iron-deficient oceanic regions. The observation that iron associated with bacterial EPS is bioavailable to eukaryotic phytoplankton (this study) could benefit bacteria indirectly by favoring eukaryotic phytoplankton growth and release of exudates, important labile substrates for heterotrophic bacteria (41). Such mutualism between eukaryotic phytoplankton and heterotrophic bacteria is probably complex as it will affect both iron chemistry and bioavailability (3).
Comparison of the chemical composition of ultrafiltered dissolved organic carbon (UDOC) in surface seawater and from eukaryotic phytoplankton isolates suggests that these microorganisms significantly contribute to the pool of EPS and uronic acids found in marine systems (24, 27, 28, 36). The bioavailability of iron associated with EPS from eukaryotic phytoplankton requires further study. EPS or UDOC produced by eukaryotic phytoplankton isolates contained 32–74% saccharides with uronic acids being the dominant fraction (28–48%; refs. 28, 36). Given that uronic acids are a major component of phytoplankton EPS, and that uronic acids enhance iron bioavailability to eukaryotic phytoplankton (this study, ref. 38), it is expected that phytoplankton EPS would have a similar effect on iron biogeochemistry to that of the saccharides studied here.
Previous reports have observed that organic matter can increase iron solubility (14), often forming colloidal organic iron (42). Iron associations formed with saccharides used in our study stabilize iron in the dissolved phase (soluble and colloidal) and are readily bioavailable to eukaryotic phytoplankton. Among the saccharides studied, only the iron associated with the EPS was able to reduce iron bioavailability (based on kupt) to the Antarctic isolates. However, iron bound to EPS was still readily bioavailable as compared with DFB-bound iron; DFB is usually reported to decrease iron bioavailability by >90% (this study, ref. 38). Hence, saccharides will increase the retention time of bioavailable forms of iron in the euphotic zone of HNLC oceanic regions, and consequently may increase the iron uptake rate by eukaryotic phytoplankton. Therefore, saccharides play a critical role in maintaining a pool of bioavailable iron in surface waters. This study demonstrates that they assist in reducing iron limitation for natural eukaryotic phytoplankton communities, diatoms and Phaeocystis, which are major contributors to downward carbon export (35, 43).
Among the saccharides studied, only the EPS formed a complex with iron that was strong enough to be detected by CLE–AdCSV, indicating that some saccharides could account for the weak ligand class (L2) measured by CLE–AdCSV (17). The formation of weak iron–saccharide associations results in an increased pool of labile iron, which are more easily exchangeable with the high-affinity iron transporters used by eukaryotic phytoplankton (9). In addition, it was shown that saccharides can favor iron reduction (33, 34) and eukaryotic phytoplankton iron uptake (this study, ref. 38), probably by the formation, via iron(II), of highly bioavailable iron species.
Our study shows that, in the iron-deficient open ocean, saccharides can significantly contribute to the operationally defined pool of SRFA-like substances detected by the technique developed by Laglera and van den Berg (40). In offshore pelagic waters, the nature of organic substances detected by this technique would be quite different from SRFA or coastal organic material, likely closer to the bacterial EPS used here. Not surprisingly, the concentrations of SRFA-equivalent substances in the euphotic zone of the Southern Ocean (this study) were much lower than for the Irish Sea and at depth in the Pacific Ocean (36–370 ppb of SRFA equivalents; refs. 16, 40). In our study, the SRFA-equivalent concentrations detected, would correspond to 4 nM of EPS in the PF and 20 nM of EPS in the SAZ.
In oceanic waters, several organic ligands are present and compete to bind iron (Fig. S1). Iron uptake by bacterioplankton (via siderophores) is specific; these microorganisms will not detect polysaccharide-bound iron, unless the iron is relocated to the siderophore via competitive ligand exchange (7). The competitive ligand exchange of iron from one organic ligand (L) to another (X) depends on their apparent affinities for iron (44). Apparent affinity is defined by the product of the stability constant and ligand concentration. In this case, iron will exchange from L to X when the apparent affinity for iron binding of the ligand X is greater than the apparent affinity for the ligand L (44). Therefore, a weak ligand can alter iron chemical speciation if present at a concentration sufficiently high to induce the iron exchange from the strong (L1) ligands.
The range of siderophores (3–20 pM; ref. 21) and saccharides (up to 500 nM, ref. 22) reported in the open ocean suggests that saccharides could induce a ligand exchange for the iron bound to siderophores. Indeed, it was shown that iron bound to domoic acid could dominate iron chemistry during a coastal bloom of a toxic Pseudonitzschia species (20). Knowing (i) the concentrations of strong organic ligands (L1) associated with iron, (ii) their stability constants measured in the SAZ and PF at the depth of chlorophyll a maximum (Table S1), and (iii) the reported stability constants for small saccharides (log KFe′L 8.8 M−1 for gluconic acid, ref. 17), one can calculate a threshold concentration of saccharides required to significantly alter iron chemical speciation and favor Fe–saccharide association. In this case, at least 206 nM of saccharides in the PF and 750 nM in the SAZ are required. Similar high concentrations of saccharides have been previously reported in the Southern Ocean (e.g., 280 nM in the Polar Front zone and 500 nM in the Ross Sea; ref. 22).
The importance of saccharides is universal, because they are present in all aquatic systems, can bind other essential micronutrients (e.g., Zn, Co, and Cu), and contain macronutrients (e.g., N as amino groups). In our study, we have demonstrated that knowledge of the source and nature of saccharides is required to understand the mechanisms of iron cycling in the Southern Ocean. The hypothesis that eukaryotic phytoplankton are able to modulate their EPS production rate and/or EPS composition in response to iron limitation requires further investigation. Whether these organisms produce saccharides to regulate iron bioavailability, as bacterioplankton produce siderophores for the same purpose, is yet to be determined.
Materials and Methods
Organic Ligands.
Four types of organic ligand (L) were chosen: (i) the strong L, DFB, a hydroxamate siderophore (Sigma-Aldrich); (ii) GLU, monosaccharide with hydroxylic and carboxylate groups (Sigma-Aldrich, 194 g mol−1); (iii) DEX, a polysaccharide, 3–6% sulfate (Sigma-Aldrich, 500,000 g mol−1); (iv) a bacterially produced EPS. The EPS was purified from a bacterium isolated from the pelagic Southern Ocean (65 °S, 143 °E), the chemical composition of which has been previously described (29). It is a large polyfunctional polysaccharide (1.7 MDa) containing mainly neutral sugars (50%), uronic acid (30%, mainly as galacturonic acid), and amino sugars (14% mainly as N-acetyl-galactosamine). The EPS presented several binding groups such as carboxylic, hydroxyl, few sulfate groups, and some associated proteins. Background iron contamination of L was checked by inductively coupled plasma mass spectroscopy (ICP-MS) (Perkin-Elmer; Elan DRC II). Iron contamination from DFB, GLU, and DEX was negligible but the EPS contained 2.15 mole of iron per mole of EPS. The C concentrations used here in perturbation experiments corresponded to 12–300 nM of C for GLU, ∼276 μM of C for DEX, and 48 μM of C for EPS. The concentrations of GLU and EPS used here were relevant to the saccharide concentrations reported in the Southern Ocean (up to 20–30 μM of C, ref. 23).
Iron Bioavailability.
Iron bioavailability, with and without the addition of various L, was measured for natural eukaryotic phytoplankton and bacterioplankton communities at two locations in the Southern Ocean and for two Antarctic eukaryotic phytoplankton isolates. The pool of iron that is bioavailable is defined here as that which can be accumulated inside microorganisms, as intracellular iron to support metabolic reactions required for their growth (Fig. S1; refs. 38, 39). On the basis of our operational definition, bioavailability is directly related to iron accumulated inside the eukaryotic phytoplankton under Fe-limited conditions. Here, iron bioavailability was measured during iron uptake experiments using a radiotracer (55FeCl3) under 50 μmol quanta m−2 s−1 for 16 h (Sun-Glo fluorescent bulb; 400–700 nm) at 2–4 °C or sea-surface temperature (Table S1). 55Iron was preequilibrated with L for at least 1 wk before experiments. Intracellular iron was determined after an EDTA-oxalate washing step, which removed iron adsorbed on the surface of the eukaryotic phytoplankton (45). Experiments in the absence of microorganisms showed that colloidal iron retention following the oxalate wash was negligible (<5% of the intracellular concentration).
Iron bioavailability was also measured using the rate constant of intracellular iron uptake (kupt, ref. 39). The kupt (h−1) was estimated by linear regression of the increase in iron uptake rate (nmol iron L−1 biovolume h−1) for five increasing total iron concentrations in synthetic seawater (nM). The kupt is dependent on both the functioning of the iron transport system (both transporter numbers and affinity for iron are important) and the water chemistry (Fig. S1; ref. 39). The kupt was measured in the presence and the absence of L for each Antarctic isolate grown in an iron-limited situation. In the absence of L and at low iron concentrations, inorganic iron is mainly present as Fe(III)′, a form that reacts directly with the transporter and surface reductase of diatoms (9); thus, it is considered here as 100% bioavailable to eukaryotic phytoplankton. Using this approach, the biological iron transport system was kept constant and only water chemistry was altered. Comparison of kupt obtained in the absence of L with kupt measured in the presence of L, allowed the estimation of iron bioavailability in the presence of L. Each experiment was performed in duplicate and Student's t test (95% significance) was used to compare iron bioavailability in the presence and absence of L.
Experiments with natural plankton were performed using Southern Ocean water collected at the depth of maximum Chl a using a trace-metal-clean protocol (46), south of the PF (54.0 °S 145.9 °E, February 1, 2007) and in the SAZ (45.6 °S 153.2 °E, February 11, 2007). Bacterioplankton (0.2–0.8 μm) and eukaryotic plankton (>0.8 μm) were distinguished using sequential filtration (polycarbonate filter; Millipore).
Antarctic eukaryotic phytoplankton isolates (diatom, Chaetoceros sp. CS-624 and haptophyte, Phaeocystis sp. CS-284) were grown in sterile Southern Ocean water (SI Materials and Methods, ref. 38), and culture axenicity was verified using DAPI staining (47). Experiments were carried out either in natural waters or in inorganic artificial seawater consisting of the major salts of the AQUIL media recipe (48) (I = 0.66 mol, pH = 8.0).
Iron Chemistry.
Chemical interaction of iron with saccharides was investigated with the CLE–AdCSV (17) (SI Materials and Methods) and sequential filtration (0.2 and 0.02 μm, Anotop; Whatman; SI Materials and Methods). Accuracy of the CLE–AdCSV method was verified using reference seawaters (NASS-5 and SAFe D2; SI Materials and Methods). In addition, operationally defined humic-like compounds were determined (40). SRFA (standard I, International Humic Substances Society) was used to calibrate the instrument (SI Materials and Methods) and concentrations were expressed in SRFA equivalents.
Acknowledgments
The authors thank Brian Griffiths and Andrew Bowie for water sampling and the opportunity to join the SAZ-Sense oceanographic campaign; the laboratory of Ken Bruland for SAFe reference seawater; Ros Watson and Jeanette O'Sullivan for ICP-MS analysis; Lesley Clementson and Corina Brussaard for HPLC pigments and flow cytometry analysis, respectively; and Tom Trull, Richard Matear, and Brian Griffiths for comments. The authors also thank the Australian Research Council for funding (DP 1092892), the Australian Antarctic Division (Australian Antarctic Science Project 2720), and the Antarctic Climate and Ecosystems Cooperative Research Center. C.S.H. was funded by the Commonwealth Scientific and Industrial Research Organization (CSIRO) Office of the Chief Executive Postdoctoral Fellowship and the University of Technology Sydney Chancellor Fellowship; C.M.N. was supported by a Postdoctoral Fellowship. The Belgian Federal Science Policy Office (Contract SD/CA/03A), Belgian French Community (Actions de Recherche Concertée Contract 2/07-287) and the Netherlands Organisation for Scientific Research (Netherlands Polar Project 851.20.046) provided financial support to V.S. The New Zealand Foundation for Research Science and Technology Coasts and Oceans Outcome Based Investment funded P.W.B. This study is also a contribution to the Surface Ocean–Lower Atmosphere Study international research initiative and the European Network of Excellence EUR-OCEANS (Contract 511106-2).
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References
1
PW Boyd, et al., Mesoscale iron enrichment experiments 1993-2005: Synthesis and future directions. Science 315, 612–617 (2007).
2
JH Martin, et al., Testing the iron hypothesis in ecosystems of the equatorial Pacific-Ocean. Nature 371, 123–129 (1994).
3
SA Amin, et al., Photolysis of iron-siderophore chelates promotes bacterial-algal mutualism. Proc Natl Acad Sci USA 106, 17071–17076 (2009).
4
DA Hutchins, AE Witter, A Butler, GW Luther, Competition among marine phytoplankton for different chelated iron species. Nature 400, 858–861 (1999).
5
MT Maldonado, RF Strzepek, S Sander, PW Boyd, Acquisition of iron bound to strong organic complexes, with different Fe binding groups and photochemical reactivities, by plankton communities in Fe-limited subantarctic waters. Global Biogeochem Cycles 19, GB4S23, 10.1029/2005GB002481. (2005).
6
RF Strzepek, et al., Spinning the “Ferrous Wheel”: The importance of the microbial community in an iron budget during the FeCycle experiment. Global Biogeochem Cycles 19, GB4S26, 10.1029/2005GB002490. (2005).
7
C Völker, DA Wolf-Gladrow, Physical limits on iron uptake mediated by siderophores or surface reductases. Mar Chem 65, 227–244 (1999).
8
AL Rose, TP Salmon, T Lukondeh, BA Neilan, TD Waite, Use of superoxide as an electron shuttle for iron acquisition by the marine cyanobacterium Lyngbya majuscula. Environ Sci Technol 39, 3708–3715 (2005).
9
Y Shaked, AB Kustka, FMM Morel, A general kinetic model for iron acquisition by eukaryotic phytoplankton. Limnol Oceanogr 50, 872–882 (2005).
10
WG Sunda, SA Huntsman, Iron uptake and growth limitation in oceanic and coastal phytoplankton. Mar Chem 50, 189–206 (1995).
11
S Wilhelm, Ecology of iron-limited cyanobacteria: A review of physiological responses and implications for aquatic systems. Aquat Microb Ecol 9, 295–303 (1995).
12
KA Hunter, PW Boyd, Iron-binding ligands and their role in the ocean biogeochemistry of iron. Environ Chem 4, 221–232 (2007).
13
EL Rue, KW Bruland, Complexation of iron(III) by natural organic ligands in the Central North Pacific as determined by a new competitive ligand equilibration/adsorptive cathodic stripping voltammetric method. Mar Chem 50, 117–138 (1995).
14
M Chen, W-X Wang, L Guo, Phase partitioning and solubility of iron in natural seawater controlled by dissolved organic matter. Global Biogeochem Cycles 18, GB4013, doi 4010.1029/2003GB002160. (2004).
15
A Tagliabue, KR Arrigo, Processes governing the supply of iron to phytoplankton in stratified seas. J Geophys Res 111, C06019 (2006).
16
LM Laglera, CMG van den Berg, Evidence for geochemical control of iron by humic substances in seawater. Limnol Oceanogr 54, 610–619 (2009).
17
P Croot, M Johansson, Determination of iron speciation by cathodic stripping voltammetry in seawater using the competing ligand 2-(2-thiazolylazo)-p-cresol (TAC). Electroanal 12, 565–576 (2000).
18
P Verdugo, et al., The oceanic gel-phase: A bridge in the DOM-POM continuum. Mar Chem 92, 67–85 (2004).
19
B Gyurcsik, L Nagy, Carbohydrates as ligands: Coordination equilibria and structure of the metal complexes. Coord Chem Rev 203, 81–149 (2000).
20
E Rue, K Bruland, Domoic acid binds iron and copper: A possible role for the toxin produced by the marine diatom Pseudo-nitzschia. Mar Chem 76, 127–134 (2001).
21
E Mawji, et al., Hydroxamate siderophores: Occurrence and importance in the Atlantic Ocean. Environ Sci Technol 42, 8675–8680 (2008).
22
C Panagiotopoulos, R Sempere, Analytical methods for the determination of sugars in marine samples: A historical perspective and futures directions. Limnol Oceanogr Methods 3, 419–454 (2005).
23
DJ Pakulski, R Benner, Abundance and distribution of carbohydrates in the ocean. Limnol Oceanogr 39, 930–940 (1994).
24
L Aluwihare, D Repeta, R Chen, A major biopolymeric component to dissolved organic carbon in surface seawater. Nature 387, 166–169 (1997).
25
AW Decho, Microbial exopolymer secretions in ocean environments: Their role(s) in food webs and marine processes. Oceanography and Marine Biology Annual Review, ed M Barnes (Aberdeen Univ Press, Aberdeen, Scotland), pp. 73–153 (1990).
26
KD Hoagland, JR Rosowski, MR Gretz, SC Roemer, Diatom extracellular polymeric substances: Function, fine structure, chemistry and physiology. J Phycol 29, 537–566 (1993).
27
I Janse, et al., Carbohydrates in the North Sea during spring blooms of Phaeocystis. Aquat Microb Ecol 10, 97–103 (1996).
28
A Biersmith, R Benner, Carbohydrates in phytoplankton and freshly produced dissolved organic matter. Mar Chem 63, 131–144 (1998).
29
CM Nichols, et al., Chemical characterization of exopolysaccharides from Antarctic marine bacteria. Microb Ecol 49, 578–589 (2005).
30
T van Oijen, et al., Enhanced carbohydrate production by Southern Ocean phytoplankton in response to in situ iron fertilization. Mar Chem 93, 33–52 (2005).
31
PH Santschi, et al., Control of acid polysaccharide production and 234Th and POC export fluxes by marine organisms. Geophys Res Lett 30, 1044, doi:1010.1029/2002GL016046. (2003).
32
MT Maldonado, MP Hughes, EL Rue, ML Wells, The effect of Fe and Cu on growth and domoic acid production by Pseudo-nitzschia multiseries and Pseudo-nitzschia australis. Limnol Oceanogr 47, 515–526 (2002).
33
M Öztürk, et al., Iron enrichment and photoreduction of iron under UV and PAR in the presence of hydroxycarboxylic acid: Implications for phytoplankton growth in the Southern Ocean. Deep Sea Res Part II Top Stud Oceanogr 51, 2841–2856 (2004).
34
S Steigenberger, PJ Statham, C Völker, U Passow, The role of polysaccharides and diatom exudates in the redox cycling of Fe and the photoproduction of hydrogen peroxide in coastal seawaters. Biogeosciences 7, 109–119 (2010).
35
V Schoemann, R Wollast, L Chou, C Lancelot, Effects of photosynthesis on the accumulation of Mn and Fe by Phaeocystis colonies. Limnol Oceanogr 46, 1065–1076 (2001).
36
S Zhang, C Xu, PH Santschi, Chemical composition and 234Th (IV) binding of extracellular polymeric substances (EPS) produced by the marine diatom Amphora sp. Mar Chem 112, 81–92 (2008).
37
KJ Sreeram, H Yamini Shrivastava, BU Nair, Studies on the nature of interaction of iron(III) with alginates. Biochim Biophys Acta 1670, 121–125 (2004).
38
CS Hassler, V Schoemann, Bioavailability of organically bound iron in controlling Fe to model phytoplankton of the Southern Ocean. Biogeosciences 6, 2281–2296 (2009).
39
I Worms, DF Simon, CS Hassler, KJ Wilkinson, Bioavailability of trace metals to aquatic microorganisms: Importance of chemical, biological and physical processes on biouptake. Biochimie 88, 1721–1731 (2006).
40
LM Laglera, G Battaglia, CMG van den Berg, Determination of humic substances in natural waters by cathodic stripping voltammetry of their complexes with iron. Anal Chim Acta 599, 58–66 (2007).
41
R Stocker, JR Seymour, A Samadani, DE Hunt, MF Polz, Rapid chemotactic response enables marine bacteria to exploit ephemeral microscale nutrient patches. Proc Natl Acad Sci USA 105, 4209–4214 (2008).
42
JT Cullen, BA Bergquist, JW Moffett, Thermodynamic characterization of the partitioning of iron between soluble and colloidal species in the Atlantic Ocean. Mar Chem 98, 295–303 (2006).
43
P Boyd, P Newton, Does planktonic community structure determine downward particulate organic carbon flux in different oceanic provinces? Deep Sea Res Part I Oceanogr Res Pap 46, 63–91 (1999).
44
FMM Morel, JG Hering Principles and Applications of Aquatic Chemistry (Wiley Interscience, New York, 1993).
45
CS Hassler, V Schoemann, Discriminating between intra- and extracellular metals using chemical extractions: An update on the case of iron. Limnol Oceanogr Methods 7, 479–489 (2009).
46
AR Bowie, MC Lohan Practical Guidelines for the Analysis of Seawater, ed O Wurl (Taylor and Francis, Boca Raton, FL), pp. 235–257 (2009).
47
KG Porter, YS Feig, The use of DAPI for identifying and counting aquatic microflora. Limnol Oceanogr 25, 943–948 (1980).
48
FMM Morel, JJG Rueter, DM Anderson, RRL Guillard, AQUIL: A chemically defined phytoplankton culture medium for trace metal studies. J Phycol 15, 135–141 (1979).
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Published online: December 15, 2010
Published in issue: January 18, 2011
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Acknowledgments
The authors thank Brian Griffiths and Andrew Bowie for water sampling and the opportunity to join the SAZ-Sense oceanographic campaign; the laboratory of Ken Bruland for SAFe reference seawater; Ros Watson and Jeanette O'Sullivan for ICP-MS analysis; Lesley Clementson and Corina Brussaard for HPLC pigments and flow cytometry analysis, respectively; and Tom Trull, Richard Matear, and Brian Griffiths for comments. The authors also thank the Australian Research Council for funding (DP 1092892), the Australian Antarctic Division (Australian Antarctic Science Project 2720), and the Antarctic Climate and Ecosystems Cooperative Research Center. C.S.H. was funded by the Commonwealth Scientific and Industrial Research Organization (CSIRO) Office of the Chief Executive Postdoctoral Fellowship and the University of Technology Sydney Chancellor Fellowship; C.M.N. was supported by a Postdoctoral Fellowship. The Belgian Federal Science Policy Office (Contract SD/CA/03A), Belgian French Community (Actions de Recherche Concertée Contract 2/07-287) and the Netherlands Organisation for Scientific Research (Netherlands Polar Project 851.20.046) provided financial support to V.S. The New Zealand Foundation for Research Science and Technology Coasts and Oceans Outcome Based Investment funded P.W.B. This study is also a contribution to the Surface Ocean–Lower Atmosphere Study international research initiative and the European Network of Excellence EUR-OCEANS (Contract 511106-2).
Notes
This article is a PNAS Direct Submission.
See Commentary on page 893.
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The authors declare no conflict of interest.
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Saccharides enhance iron bioavailability to Southern Ocean phytoplankton, Proc. Natl. Acad. Sci. U.S.A.
108 (3) 1076-1081,
https://doi.org/10.1073/pnas.1010963108
(2011).
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